Direct and indirect functions are assigned to von Willebrand Factor in normally proceeding blood coagulation. It binds to Factor VIII in a complex. This complex serves to stabilize Factor VIII. This stabilized Factor VIII then has essential cofactor function in the activation of Factor X. Additionally, von Willebrand Factor directly influences blood coagulation by mediating platelet aggregation to injured vessels.
In plasma, vWF circulates in a concentration of 5-10 mg/l in the form of a non-covalent complex with Factor VIII. vWF is a glycoprotein which is formed in different cells of the human body and is later released into the circulation. Moreover, starting from a polypeptide chain with a molecular weight of about 220,000 (vWF monomer), a vWF diner (primary diner) with a molecular weight of about 550,000 is made in cells by the formation of several sulfur bridges. Then, further polymers of vWF with increasing molecular weights up to about 20 million are produced from the vWF dimers by association.
There are several clinical pictures which are traceable to under- or overproduction of von Willebrand Factor. Thus, for example, an overproduction of vWF leads to an increased tendency towards thromboses, whereas an undersupply of vWF results in an increased bleeding tendency or prolonged bleeding time.
von Willebrand Syndrome can manifest itself in several forms. All forms are distinguished by a prolonged bleeding time which is based on either an absolute absence of a functional vWF or an abnormal spectrum in the multimer composition of vWF. Forms of von Willebrand disease in which multimer formation is reduced as well as forms in which low molecular vWF molecules are barely present are diagnosed thereby. Although other forms demonstrate high and low molecular vWF molecules, their concentration and/or their ratio to each other is drastically decreased and/or altered compared to a healthy person.
The lack of vWF can also cause hemophilia A because vWF, as mentioned above, is an essential component of functional Factor VIII. In these cases, the half-life of Factor VIII is decreased in such a manner that it can not fulfill its special functions in the blood coagulation cascade.
All forms of von Willebrand Syndrome as well as the form of hemophilia traceable to the lack of vWF were treated up to now by replacement of the missing vWF through intravenous infusions with concentrates of blood plasma which contain either vWF-Factor VIII complex or enriched vWF. Although in one respect the administration of Factor VIII is not necessary in both disease cases, the preparative separation of vWF from Factor VIII is technically very difficult to impossible.
In order to establish the exact function of high molecular vWF molecules on the one hand and low molecular vWF molecules on the other hand, ways have been looked for to isolate these fractions in enriched form. Wagner, D. D. et al (J. Cell Biol. 101: 112, 1985) prevent multimer formation by addition of monensin in vitro and conclude from different experiments that the low molecular form of vWF is not functional. On the other hand, Senogles, S. E. et al (J. Biol. Chem. 258: 12327, 1983) find no functional difference in ristocitin mediated platelet aggregation in high molecular and low molecular forms of vWF. They obtain low molecular forms by reduction of the sulfur bridges of high molecular vWF molecules. Aihara, M. et al (Tohoku J. Exp. Med. 153: 169, 1985) observe a lower binding capacity of vWF molecules from patients with von Willebrand Syndrome of the type IIa to collagen in affinity chromatography. This type of von Willebrand Syndrome is characterized by absence of the high molecular molecules of vWF.
In the literature, there are numerous methods which describe an analytical separation of high molecular and low molecular forms of von Willebrand Factor such that statements can be made as to the quantitative proportion of both forms. As an example, a publication from Baillod et al in Thrombosis Res. 66: 745, 1992 should be mentioned here. However, a preparative method for the separation of low molecular and high molecular forms has not been described to date.